Measuring cis-regulatory energetics in living cells using allelic manifolds

Forcier, T. L., Ayaz, A., Gill, M. S., Jones, D., Phillips, R., Kinney, J. B. (December 2018) Measuring cis-regulatory energetics in living cells using allelic manifolds. eLife, 7. ISSN 2050084X (ISSN)

[thumbnail of 2019.Forcier.manifolds.pdf]
Preview
PDF
2019.Forcier.manifolds.pdf - Published Version
Available under License Creative Commons Attribution.

Download (2MB) | Preview
URL: https://www.ncbi.nlm.nih.gov/pubmed/30570483
DOI: 10.7554/eLife.40618

Abstract

Gene expression in all organisms is controlled by cooperative interactions between DNA-bound transcription factors (TFs), but quantitatively measuring TF-DNA and TF-TF interactions remains difficult. Here we introduce a strategy for precisely measuring the Gibbs free energy of such interactions in living cells. This strategy centers on the measurement and modeling of 'allelic manifolds', a multidimensional generalization of the classical genetics concept of allelic series. Allelic manifolds are measured using reporter assays performed on strategically designed cis-regulatory sequences. Quantitative biophysical models are then fit to the resulting data. We used this strategy to study regulation by two Escherichia coli TFs, CRP and sigma-70 RNA polymerase. Doing so, we consistently obtained energetic measurements precise to ~0.1 kcal/mol. We also obtained multiple results that deviate from the prior literature. Our strategy is compatible with massively parallel reporter assays in both prokaryotes and eukaryotes, and should therefore be highly scalable and broadly applicable. Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that minor issues remain unresolved (see decision letter).

Item Type: Paper
Subjects: bioinformatics
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification
bioinformatics > genomics and proteomics > genetics & nucleic acid processing
bioinformatics > genomics and proteomics
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > genes, structure and function > alleles
organism description > bacteria
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > cis-regulatory elements
organism description > bacteria > escherichia coli
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > genes, structure and function
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types
bioinformatics > genomics and proteomics > genetics & nucleic acid processing > protein structure, function, modification > protein types > transcription factor
CSHL Authors:
Communities: CSHL Cancer Center Program > Gene Regulation and Inheritance Program
CSHL labs > Kinney lab
School of Biological Sciences > Publications
Depositing User: Matthew Dunn
Date: 20 December 2018
Date Deposited: 07 Jan 2019 15:49
Last Modified: 29 Feb 2024 20:32
PMCID: PMC6301791
Related URLs:
URI: https://repository.cshl.edu/id/eprint/37529

Actions (login required)

Administrator's edit/view item Administrator's edit/view item
CSHL HomeAbout CSHLResearchEducationNews & FeaturesCampus & Public EventsCareersGiving